The ability to control inflammatory and immune responses is central to the therapy of a wide spectrum of diseases. General anti-inflammatory agents (corticosteroids and non steroidal anti-inflammatory drugs, i.e. NSAIDs, such as aspirin) acting to suppress or regulate immune-mediated reactions throughout the body, are widely used in this context. The modulation of receptors and various functions of leukocytes which participate in, and are responsible for, the local response of the injured tissue, illustrates a new approach in clinical patient care. Biological agents, such as antibodies directed to inflammatory leukocytes or their receptors, provide important control mechanisms through which immune-mediated tissue destruction can be prevented.
Specific receptors with binding affinity for a variety of ligands are found on the surface of cells that participate in the inflammatory response. CD44 (also referred to as PgP-1, phagocytic glycoprotein-1) is a cell adhesion receptor which preferentially binds hyaluronan (Aruffo et al., Cell, 61 (1990) 1303-1313). CD44 is expressed by many cell types (Lesley et al., Adv. in Immunol., 54 (1993) 271-335) and expressed in high levels by synovial cells and leukocytes during joint inflammation (Haynes et al., Arthritis Rheum., 34 (1991) 1434-1443; Mikecz et al., Scand. Rheumatol., 101 (1994) 91-98). The CD44 glycoprotein is expressed in several isoforms as a result of differential splicing (Gunthert et al., Curr Topics Microbiol. Immun., 184 (1993) 47-63; Gunthert et al., Cell, 65 (1991) 13-24; MacKay et al., J.Cell Biol., 124 (1994) 71-82).
Leukocytes responsible for immune-mediated tissue injury, and cells residing in the tissue and extracellular matrix function together to effect an inflammatory response. The extracellular matrix (ECM) fills the space between cells. Although it has long been recognized that the components of the ECM perform an important structural role, it has been realized more recently that the ECM communicates with the cell interior and thus modulates cell adhesion, proliferation, and differentiation (Schubert, Trends Cell Biol., 2 (1992) 63-66). Major constituents of the ECM include collagenous proteins (Linsenmayer, Cell Biology of the Extracellular Matrix, Plenum, N.Y. (1991) 7-44) and proteoglycans. The latter consist of one or more glycosaminoglycans, which are linear polymers of repeating disaccharides covalently bound to a protean core. Hyaluronan (hyaluronic acid, hyaluronate), a glycosaminoglycan macromolecule without a protein core, is one of the major non-structural elements of the extracellular matrix (Laurent et al., FASEB J., 6 (1992) 2397-2404; Aruffo et al., Cell, 61 (1990) 1303-1313; Culty et al., J. Cell Biol., 111 (1990) 2765-2774; Underhill, J. Cell Sci., 103 (1992) 293-298; Toole, Cell Biology of Extracellular Matrix, (1991) Plenum, N.Y.).
The biological roles of hyaluronan include the maintenance of water and protein homeostasis as well as the protection of cells from potentially harmful effects of other cells, microorganisms and macromolecules. To gain access to cells surrounded by hyaluronan-rich matrices, some cells and bacteria use hyaluronidase. Other cells utilize hyaluronan binding receptors such as CD44 (Laurent et al., FASEB J., 6 (1992) 2397-2404; Toole, Cell Biology of Extracellular Matrix, (1991) Plenum, N.Y.; Lesley et al., Exp. Cell Res., 187 (1990) 224-233; Bartolazzi et al., J. Exp. Med., 180 (1994) 53-66; Thomas et al., J. Cell Biol., 118 (1992) 971-977; Herrlich et al., Immun. Today, 14 (1993) 395-399). The interaction of CD44 with hyaluronan facilitates the migration of these cells within the extracellular matrix. Further, hyaluronan molecules have large hydrodynamic volumes that entrap substantial amounts of water and can, hence, control tissue hydration (swelling).
Lymphocytes acquire enhanced binding affinity toward hyaluronan upon activation (Stamenkovic et al., Embo J., 10 (1991) 343-348; Hathcock et al., J. Immun., 151 (1993) 6712-6722). Hyaluronan-binding splice variants of CD44 confer metastatic proclivity to malignant cells (Gunthert et al., Cell, 65 (1991) 13-24). Recombinant CD44H (hemopoietic form) proteins and antibodies that recognize CD44H inhibit both cell adhesion to hyaluronan and migration of cells on hyaluronan-coated surfaces. Taken together, these findings suggest an important role for the CD44-hyaluronan interaction during physiological and pathological events in which cell migration within the ertracellular space is involved.
Researchers have attempted to modulate the immune response by developing antibodies to specific receptors and hence prevent binding of any other ligand to that receptor. A number of patents describe antibodies directed to various adhesion associated molecules. For example, U.S. Pat. No. 5,147,637 describes a method of inhibiting the influx of leukocytes into the lung and other organs during sepsis or other infectious or non-infectious trauma by administering a therapeutic amount of and anti-CD18 antibody. U.S. Pat. No. 4,695,459 describes a method of treating autoimmune disease, such as multiple sclerosis, rheumatoid arthritis and systemic lupus erythematosus by administering a therapeutically effective amount of anti-Leu3 antibody to the patient, which would eliminate T lymphocytes. Further, U.S. Pat. No. 5,019,648 describes the use of anti-CD11b antibodies to inhibit adhesion dependent functions of phagocytic cells. None of these patents describe CD44 receptor, its interaction with hyaluronan or a method wherein antibody binding results in the loss of receptors from the cell surface.
Methods have been described for the therapeutic use of antibodies to prevent leukocyte extravasation in individuals having autoimmune diseases such as rheumatoid arthritis. For example, U.S. Pat. No. 5,216,131 describes the therapeutic use of lymphocyte homing receptor protein (LHR) to compete with the normal binding of lymphocytes to lymphoid tissue for the treatment of patients with inflammations due to rheumatoid arthritis or other autoimmune diseases. European Patent Application 303,463 and corresponding U.S. Pat. No. 5,403,919 describe antibodies having recognition for a 58-69 kD mouse endothelial cell surface antigen present on the surface of high endothelial venule (HEV) cells in all lymphoid organs, and a 85-95 kD synovial-specific leukocyte glycoprotein which is a homing receptor for synovial endothelium. The antibodies having recognition for these endothelial cells surface antigens are capable of inhibiting the binding of leukocytes to endothelial cells, thereby inhibiting lymphocyte extravasation via such endothelial cells in vivo.
The antigens described in 5,216,131 and EP 303,463 are structurally and functionally different from CD44. CD44 is not a homing receptor as none of the isoforms of CD44 contain binding sites for endothelial cell-membrane carbohydrates which are commonly recognized by homing receptors, and anti-CD44 antibodies do not disturb normal lymphocyte homing in vivo (Camp et al., J. Exp. Med., 178 (1993) 497-507; Mikecz et al., Nature Med., 1 (1995) 558-563). Further, CD44 is the principal receptor for hyaluronan, which is a component of extracellular or pericellular matrix, and the digestion of lymphoid tissues with hyaluronidase eliminates CD44 binding. Thus, CD44-bearing leukocytes do not recognize endothelial cells directly in either the lymph node or synovial tissue. Leukocyte CD44, however, can bind hyaluronan, which is not an integral part of the endothelial cell, but may be present in the extra- or pericellular matrix around the endothelium, especially in inflamed tissue where hyaluronan is abundantly produced.
CD44 is not synovial specific leukocyte glycoprotein as it has been shown to be expressed in many cell types besides leukocytes (Lesley et al., Adv. in Immunol., 54 (1993) 271-335; Mikecz et al., Scand. J. Rheumatol., 101 (1994) 91-98). EP 303,4463 describes antibodies which are able to inhibit leukocyte binding to the endothelium of target organs by blocking leukocyte-endothelial cell recognition, i.e. the antibodies block cell-cell contact but not hyaluronan-cell interaction. Hence, none of the references describe an antibody having a twofold anti-inflammatory effect provided by binding CD44 receptors on connective tissue cells and leukocyte CD44 receptors such that receptors are lost from the cell surface, hyaluronan binding and associated swelling is reduced, and leukocyte migration is inhibited.
It is thus an object of the present invention to provide an immunotherapeutic method for the treatment of humans or animals to control autoimmune inflammatory diseases.
Another object of the invention is to provide a therapeutic regimen which provides the maximal desired anti-inflammatory effect balanced with the least adverse side effects.
Yet another object of the invention is to provide an immunotherapeutic method wherein antibodies to CD44 receptors are utilized to inhibit the interaction of CD44 receptors with hyaluronan such that tissue swelling is reduced and leukocyte extravasation is inhibited.
It is an additional object of the invention to provide a method wherein the binding of antibodies to CD44 results in the loss of CD44 receptor from the surface of the cell.